Efficient vaccine production requires the growth of large scale quantities of virus produced in high yields from a host system. The cultivation conditions under which a virus strain is grown is of great significance with respect to achieving an acceptable high yield of the strain. Thus, in order to maximize the yield of the desired virus, both the system and the cultivation conditions must be adapted specifically to provide an environment that is advantageous for the production of the desired virus. Therefore, in order to achieve an acceptably high yield of the various virus strains, a system which provides optimum growth conditions for a large number of different virus is required.
The only process which is economically viable is a reactor process because the scale-up can be made appropriate to the market size and the vaccine doses needed. For adherent cells the carrier process with a classical microcarrier is currently the best choice for large scale cultivation of the cells needed for virus propagation (Van Wezel et al. 1967. Nature 216:64-65; Van Wezel et al. 1978. Process Biochem. 3:6-8). Large-scale process production of poliomyelitis virus, Hepatitis A Virus, HSV or Mareck's disease virus on microcarrier has been described (U.S. Pat. No. 4,525,349; Widell et al., 1984. J. Virological Meth. 8:63-71; Fiorentine et al., 1985. Develop. Biol. Standard 60:421-430; Griffiths et al., 1982. Develop. Biol. Standard. 50:103-110). Current processes based on microcarrier culture allow production of virus using fermenter sizes of up to 1200 l.
Caij et al. (1989. Arch. Virol. 105: 113-118) compared production yields of virus titre of Hog Cholera Virus on microcarrier cultures and conventional monolayer cultures and found that using the microcarrier system higher virus yield per volume of medium can be obtained.
Griffiths et al. (1982. Develop. Biol. Standard. 50:103-110) studied the influence of the microcarrier concentration on cell growth and production of HSV. It was found that an optimal concentration of microcarriers is needed to reach high cell density, which also influences the virus yield obtained. Higher concentrations of microcarrier in a perfusion system, however, resulted in a cell loss due to cell layer sloughing off the beads.
The productivity of the virus production process on the microcarrier system depends on the virus, the cells, the type of microcarrier and the cell density obtained in the system. Higher microcarrier concentrations in the cell culture allow for higher total cell numbers. However, microcarriers are costly and, in these conditions, cell loss may occur due to the cell layers sloughing off the beads by the shearing force in the system. This implies that for higher virus yields a larger volume of microcarrier cell culture is needed, but this increases the efforts that have to be made for processing and purification such large volumes.
For virus propagation it is important that optimal cell density is reached to obtain maximal virus yield. It is also important to allow efficient adsorption of virus to the cells. In conventional methods, therefore, the volume of the growth medium is reduced prior to infection to allow adsorption of the virus to the cells in a minimum of culture volume and for a better virus to cells ratio. However, to obtain optimal virus propagation, the culture medium volume is again increased after appropriate adsorption time to allow the cells to maintain viability and/or growth. This, however, increases the culture medium volume comprising cells and/or virus which has the disadvantage that large volumes have to be processed for further purification of the virus from the cells or the cell culture medium.
In the case of an outbreak of a virus infection, it is critical to produce large amounts of a vaccine in a timely fashion to provide several million vaccine doses within a very short period of time. Therefore, a continuing need exists for safe and effective methods to produce viruses and antigens. Moreover, there is a need for an approach to viral propagation, employing materials that are already available and requiring a minimal number of time-consuming manipulations, such as handling of reduced volumes of cell culture medium and facilitate purification and down stream processing for vaccine production.